TaqMan-Technology/Expression Profiling
TaqMan is a recently developed technique, in which the release of a fluorescent reporter dye from a hybridisation probe in real-time during a polymerase chain reaction (PCR) is proportional to the accumulation of the PCR product. Quantification is based on the early, linear part of the reaction, and by determining the threshold cycle (CT), at which fluorescence above background is first detected.
Gene expression technologies may be useful in several areas of drug discovery and development, such as target identification, lead optimization, and identification of mechanisms of action. The TaqMan technology can be used to compare differences between expression profiles of normal tissue and diseased tissue. Expression profiling has been used in identifying genes, which are up- or downregulated in a variety of diseases. An interesting application of expression profiling is temporal monitoring of changes in gene expression during disease progression and drug treatment or in patients versus healthy individuals. The premise in this approach is that changes in pattern of gene expression in response to physiological or environmental stimuli (e.g., drugs) may serve as indirect clues about disease-causing genes or drug targets. Moreover, the effects of drugs with established efficacy on global gene expression patterns may provide a guidepost, or a genetic signature, against which a new drug candidate can be compared.
LTBP2
The nucleotide sequence of LTBP2 is accessible in the databases by the accession number Z37976 (human) and Y12760 (rat). The sequences are given in SEQ ID NO:1 (human) and SEQ ID NO:2 (rat). The amino acid sequence of LTBP2 depicted in SEQ ID NO:3 (human) and SEQ ID NO:4 (rat).
The transforming growth factor beta (TGFβ) cytokines are a multifunctional family that exert a wide variety of effects on both normal and transformed mammalian cells. The secretion and activation of TGFβs is regulated by their association with latency associated proteins and latent TGFβ binding proteins (LTBPs). Transforming growth factor β (TGFβ) exists as three mammalian isoforms (TGFβ1, TGFβ2 and TGFβ3). Each of these is usually secreted in large latent complexes (LLCs) which have no biological activity and comprise three components: a disulphide bonded homodimer of mature TGFβ associated non-covalently with latency-associated proteins (LAPs; homodimers of the N-terminal fragment of precursor TGFβ) and a covalently attached molecule of latent TGFβ binding protein (LTBP) Four LTBP genes have been identified: LTBP1 to LTBP4. LAPs are sufficient to render the mature homodimer inactive, and removal of both the LAPs and LTBP or modulation of their interaction is essential for any of the TGFβ isoforms to function. The TGFβ cytokines modulate the growth and functions of a wide variety of mammalian cell types. It has become evident in recent years that LTBPs may be involved in the assembly, secretion and targeting of TGFβ to sites at which it is stored and/or activated. Thus these proteins may play critical roles in controlling and directing the activity of TGFβs. LTBPs may also exert effects independently of those associated with TGFβ, for example as structural matrix proteins [Oklu et al., (2000)]
Relatively little is known about the functional role of LTBP2. Unlike the other LTBPs, LTBP2 is unable to associate with the small latent TGFβ [Saharinen et al. (2000)] Human LTBP2 is expressed mostly in the lung and to a lesser extent in the liver, skeletal muscle placenta and heart [Vehvilainen et al (2003)]. Latent TGFβ binding protein LTBP2 decreases fibroblast adhesion to fibronectin [Hyytiainen et al. (2003)] Elucidation of the functional role of LTBP2 is further limited by the fact that deletion of LTBP2 in mice leads to embryonic lethality [Shipley et al. (2000)].
Regarding a functional role of LTBP2 in the cardiovascular system, it was demonstrated that LTBP2 synthesis increased in response to arterial injury in a porcine model of coronary angioplasty [Sinha et al. (2000)] Thus, together with the well known role of TGFβ in the developing of heart failure [Watkins et al. (2006)] our finding that TGFβ-function modifying LTBP2 is regulated on RNA level in LVAD hearts as well as in various animal models of heart failure makes LTBP2 an attractive candidate biomarker for CHF.
LTBP2 is published (but not limited to) in patents WO2004075835 and WO02068579.